The H9c2(2-1) cell-based sulforhodamine B assay is a non-animal alternative to evaluate municipal wastewater quality over time

Abstract

In the context of alternatives to animal testing, and since wastewater composition is highly variable, depending on time, day of the week and season, the present study validates the potential of the H9c2(2 − 1) cell-based sulforhodamine B (SRB) assay to evaluate the temporal variability of municipal wastewater. The impact of effluent disposal on water quality was also assessed using indicators of possible nutrient enrichment in the receiving water, and by determining ammonium, pH, chemical oxygen demand, total suspended solids and nine metal elements. Since rainwater can enter the municipal wastewater collection system and possibly alter toxicity, precipitation data was considered, as well as the number of new Covid-19 cases per week in the municipality. Moreover, the efficiency of the wastewater treatment process was evaluated as both raw influent and treated effluent samples were analysed. Results revealed that the H9c2(2 − 1) cell-based SRB assay appears to have an enormous potential to evaluate the temporal variability of municipal effluents, to discriminate influent and effluent toxic characteristics, as well as to study Covid-19 progression. We were able to trace the development of this pandemic by relating positive rates (wastewater may also reflect asymptomatic individuals), a crucial metric for understanding the development of a pandemic, with the selected alternative in vitro platform. Finally, the gathered results alerted to the impact of phosphates in the aquatic environment, leading us to recommend the inclusion of this parameter in regulatory assessments of wastewater discharges.